Semi-continuous Bioreactor Research Articles

Valorization of crude glycerol and eggshell biowaste as media components for hydrogen production: A scale-up study using co-culture system

The properties of eggshells (EGS) as neutralizing and immobilizing agent were investigated for hydrogen (H2) production using crude glycerol (CG) by co-culture system. Eggshells of different sizes and concentrations were used during batch and repeated-batch fermentation. For batch and repeated-batch fermentation, the maximum H2 production (36.53±0.53 and 41.16±0.95mmol/L, respectively) was obtained with the EGS size of 33μm<x5<75μm. Hydrogen production increased with the decreasing size of EGS. Eggshells maintained the fermentation pH (6.00–6.30) and provided immobilization support as confirmed by scanning electron microscopy. As media components, the EGS concentration of 0.25% (w/v) was found to be optimum for maximum H2 production (31.66±0.55mmol/L) and the production profile was comparable to H2 production (32.07±0.92mmol/L) obtained with all media components. In scale-up study with semi-continuous bioreactor (7.5L), almost 1.5-fold increase (in comparison to mono-culture) i.e. 312.12mmol-H2/L-of medium with 86.65% glycerol utilization was obtained.

Acclimation strategy to increase phenol tolerance of an anaerobic microbiota

A wide variety of inhibitory substances can induce anaerobic digester upset or failure. In this work the possibility to improve the resistance of an anaerobic microbiota to a common pollutant, the phenol, was evaluated in a lab-scale semi-continuous bioreactor. An acclimation strategy, consisting in a regular step-wise adaptation of the microbiota to stressful condition was employed. Degradation performances were monitored and molecular tools (16S sequencing and ARISA fingerprinting technique) were used to track changes in the microbial community. The acclimation strategy progressively minimized the effect of phenol on degradation performances. After 3 successive disturbance episodes, microbiota resistance was considerably developed and total inhibition threshold increased from 895 to 1942mg/L of phenol. Microbiota adaptation was characterized by the selection of the most resistant Archaea OTU from Methanobacterium genus and an important elasticity of Bacteria, especially within Clostridiales and Bacteroidales orders, that probably enabled the adaptation to more and more stressful conditions.

Semicontinuous Bioreactor Production of Recombinant Butyrylcholinesterase in Transgenic Rice Cell Suspension Cultures.

An active and tetrameric form of recombinant butyrylcholinesterase (BChE), a large and complex human enzyme, was produced via semicontinuous operation in a transgenic rice cell suspension culture. After transformation of rice callus and screening of transformants, the cultures were scaled up from culture flask to a lab scale bioreactor. The bioreactor was operated through two phases each of growth and expression. The cells were able to produce BChE during both expression phases, with a maximum yield of 1.6 mg BChE/L of culture during the second expression phase. Cells successfully regrew during a 5-day growth phase. A combination of activity assays and Western blot analysis indicated production of an active and fully assembled tetramer of BChE.

Digestión anaeróbica de desechos de frutas y hortalizas en reactores semicontinuos de un mercado municipal en Tumbaco, Ecuador

En Ecuador se generan una gran cantidad de residuos orgánicos. En el año 2010, el promedio de recolección de desechos en el Distrito Metropolitano de Quito fue de aproximadamente 1500 t por día, de los cuales más del 60% fueron identificados como residuos orgánicos. Los rellenos sanitarios que son comúnmente empleados en el país para el tratamiento de desechos generan grandes problemas al medio ambiente y a la salud pública. El tratamiento biológico de residuos orgánicos bajo condiciones anaerobias constituye una alternativa viable para estabilizar la materia orgánica, proteger el medio ambiente y reducir el riesgo de afección a la salud pública; adicionalmente, se generan productos con valor agregado como son el biogás y el fertilizante orgánico que pueden ser empleados como fuente de energía alternativa y en las actividades agrícolas, respectivamente. En el presente trabajo de investigación, el tratamiento anaeróbico de residuos sólidos de frutas y vegetales (RFV) del mercado central de Tumbaco fue estudiado en cuatro biorreactores de 4L en modo de operación semi-continuo escala laboratorio. En primer lugar los RFV fueron caracterizados para ser utilizados como materia prima en el proceso de digestión anaeróbica. A continuación, los biorreactores fueron incubados con 10% de inóculo microbiano preparado y diferentes porcentajes en peso de residuos de frutas y vegetales con el fin de identificar la concentración optima de carga orgánica. La composición del inoculo microbiano preparado fue 40% de lodos granulares provenientes del biodigestor de una planta de tratamiento de aguas de la ciudad de Quito y 90% (v/v) de estiércol vacuno disuelto. Los biodigestores mesófilos de mezcla completa fueron alimentaron con diferentes porcentajes en peso de RFV equivalentes a 5, 7.5, 10 and 12.5% sólidos totales. Los bioreactores fueron operados durante 90 días en tres períodos correspondientes a las diferentes alimentaciones. La mayor produccion acumulada de metano se alcanzó con el biorreactor alimentado con 5% RFV registrando un valor de 2401.90 L CH4 kg SV-1. Finalmente, una tecnología efectiva de digestión anaerobia fue exitosamente desarrollada como alternativa a los rellenos sanitarios para el tratamiento de residuos orgánicos municipales en Ecuador.

Biological pretreatment of non-flocculated sludge augments the biogas production in the anaerobic digestion of the pretreated waste activated sludge

High-efficiency resource recovery from municipal solid waste (MSW) has been a focus of attention. The objective of this research is to develop a bio-pretreatment process for application prior to the anaerobic digestion of MSW to improve methane productivity. Bacillus licheniformis was used for pretreating MSW (non-flocculated with 0.07% citric acid), followed by anaerobic digestion. Laboratory-scale experiments were carried out in semi-continuous bioreactors, with a total volume of 5 L and working volume of 3 L. Among the nine organic loading rates (OLRs) investigated, the OLR of 0.84 kg SS m−3 reactor day−1 was found to be the most appropriate for economic operation of the reactor. Pretreatment of MSW prior to anaerobic digestion led to 55% and 64% increase of suspended solids (SS) and volatile solids reduction, respectively, with an improvement of 57% in biogas production. The results indicate that the pretreatment of non-flocculated sludge with Bacillus licheniformis which consumes less energy compared to other pretreatment techniques could be a cost-effective and environmentally sound method for producing methane from MSW.

Semicontinuous bioreactor production of a recombinant human therapeutic protein using a chemically inducible viral amplicon expression system in transgenic plant cell suspension cultures

Plant cell culture is an alternative for the production of recombinant human therapeutic proteins because of improved product safety, lower production cost, and capability for eukaryotic post-translational modification. In this study, bioreactor production of recombinant human alpha-1-antitrypsin (rAAT) glycoprotein using a chemically inducible Cucumber mosaic virus (CMV) viral amplicon expression system in transgenic Nicotiana benthamiana cell culture is presented. Optimization of a chemically inducible plant cell culture requires evaluation of effects of timing of induction (TOI) and concentration of inducer (COI) on protein productivity and protein quality (biological functionality). To determine the optimal TOI, the oxygen uptake rate (OUR) of the plant cell culture was chosen as a physiological indicator for inducing maximum rAAT expression. Effects of COI on rAAT production were investigated using a semicontinuous culture, which enables the distinction between effects of growth rate and effects of inducer concentration. An optimized semicontinuous bioreactor operation was further proposed to maximize the recombinant protein production. The results demonstrated that the transgenic plant cells, transformed with the inducible viral amplicon expression system, maintain higher OUR and exhibit lower extracellular protease activity and lower total phenolics concentration in the optimized semicontinuous bioreactor process than in a traditional batch bioreactor operation, resulting in a 25-fold increase in extracellular functional rAAT (603 microg/L) and a higher ratio of functional rAAT to total rAAT (85-90%). Surprisingly, sustained rAAT production and steady state, long-term bioreactor operation is possible following chemical induction and establishment of the viral amplicons.

Anaerobic digestion of the vinasses from the fermentation of Agave tequilana Weber to tequila: The effect of pH, temperature and hydraulic retention time on the production of hydrogen and methane

The objective of this work was to study the effect of three operational parameters (pH, hydraulic retention time (HRT) and growing temperature) on a semi-continuous bioreactor treating Tequila's vinasses by anaerobic digestion (AD). The response was measured through four response variables: total reducing sugars (TRS) consumption, VFA's, hydrogen and methane production. Trials were done according to a factorial design. The experimental results were studied through a multiple response optimization (MRO) analysis to find single and multiple optimums for the above-mentioned variables. Mathematical models that can describe the effect of the operational parameters on each response variable were found. In this study it is shown that hydrogen production is favored at thermophilic growth (55 °C), operating the reactor at a slight acidic pH range and at the higher HRT in the boundaries of the experimental region.

Shoot Micropropagation and Microtuberization in Potato (Solanum tuberosum L.) By the Semi-Continuous Bioreactor

The aim of this investigation is introducing the use of continuous and semi-continuous bioreactors and their functions at shoot multiplication and microtuberization of potato. The study shows that the explants have several nodes and when they are suspended continuously under the liquid culture medium, the shoot micropropagations and microtuberization was inhibited. The surfaces of the explants were formed callus and subsequently, they were died. However, in the semicontinuous bioreactor, with the periodical pumping of the nutritional medium the explants aren't continuously suspended under the nutritional solution. The shoot micropropagation, the leaf growth and, the root formation are suitable. In the microtuber inductive medium, the numerous of the tubers are induced. They were sessile tubers. The dormancy of the tubers are long-term, spourting after 3-4 months in the room conditions.

Adsorption and biological decolourization of azo dye Reactive Red 2 in semicontinuous anaerobic reactors

The microbial decolourization of Reactive Red 2 (RR2) dye has been studied under anaerobic conditions. Three semicontinuous bioreactors were operated with dye concentrations—R1 (control: 0 mg RR2 l −1), R2 (100 mg RR2 l −1) and R3 (200 mg RR2 l −1). The parameters monitored were, oxidation–reduction potential (ORP), methane production, colour and chemical oxygen demand (COD) removal during the feeding cycles. The oxidation–reduction potential values for the first few days were above −150 mV, which later on decreased to less than −275 mV in all the reactors. Colour removal during the first few days of operation was due to adsorption of dye on to anaerobic biomass. However, under steady state conditions, colour removal was above 76% for both the dye containing reactors and it was due to biologically mediated degradation. Methane production and chemical oxygen demand removal in the control and dye containing reactors were almost the same. Integrated analysis of the monitored parameters indicated that, the primary mechanism of colour removal was adsorption of RR2 on to anaerobic biomass and subsequent degradation. Decolourization rates were found to be first order with respect to dye concentration, although an increase in the influent dye concentration resulted in a decrease in the rate from 0.0074 (g volatile suspended solid, VSS) −1 h −1 (100 mg RR2 l −1) to 0.0039 (g VSS) −1 h −1 (200 mg RR2 l −1). Based on total methane production no inhibition effect of dyes was observed but total methanogenic activity (TMA) results exhibited inhibition of methanogenesis.

Tight control of growth and cell differentiation in photoautotrophically growing moss (Physcomitrella patens) bioreactor cultures

The use of the moss Physcomitrella patens as a production system for heterologous proteins requires highly standardised culture conditions. For this purpose a semi-continuous photoautotrophic bioreactor culture of Physcomitrella was established. This culture grew stably for 7 weeks in a 5-l bioreactor with a dilution rate of 0.22/day. Enrichment of the air for aeration in a batch bioreactor culture with 2% (v/v) CO 2 resulted in an in- crease in the specific growth rate to 0.57/day. Changes in the pH of the semi-continuous bioreactor culture medi- um between pH 4.5 and pH 7.0 influenced protonema differentiation; however it did not negatively affect the growth rate compared to uncontrolled pH. The advanta- ges of Physcomitrella as a system for the production of heterologous proteins in plants are discussed.

Optimisation of a bioreactor culture of the moss Physcomitrella patens for mass production of protoplasts

One prerequisite for plant functional genomic projects is the development of a high-throughput transformation platform. As the moss Physcomitrella patens can be transformed via PEG-mediated DNA-uptake into protoplasts, a semi-continuous bioreactor culture of this plant was optimised regarding protoplast isolation efficiencies. Under standard conditions protoplast yields were 2.8×10 4/mg dry weight. This yield was increased sixfold by supplementation of the medium with 460 mg/l ammonium tartrate. The same effect was achieved by controlling the pH-value in the bioreactor culture with a setpoint of 4.5. In contrast, pH control with a setpoint of 7.5 reduced the protoplast yield compared to a culture without pH control to 11%. A semi-continuous culture of Physcomitrella in a 5-l bioreactor grown at pH 4.5 yielded sufficient cell material for more than 100 transformations per day.

Designing robust optimal dynamic experiments

Abstract The quality of experiments designed using standard optimisation-based techniques can be adversely affected by poor starting values of the parameters. Thus, there is a necessity for design methods that are insensitive (“ robust ”) to these starting values. Here, two novel, robust criteria are presented for computing optimal dynamic inputs for experiments aiming at improving parameter precision, based on previously proposed methods for providing design robustness — the expected value criterion and the max–min criterion. In this paper, both criteria are extended by use of the information matrix derived for dynamic systems. The experiment design problem is cast as an optimal control problem, with experiment decision variables such as sampling times of response variables, time-varying and time-invariant external controls, experiment duration, and initial conditions. Comparisons are made of experiment designs obtained using the conventional approach and the newly proposed criteria. A typical semi-continuous bioreactor application is presented.

Robust Optimal Designs for Dynamic Experiments

Abstract The quality of experiments designed using standard optimisation-based techniques can be adversely affected by poor starting values of the parameters. Thus, there is a necessity for design methods that are insensitive ( “robust” ) to these starting values. Here, a novel, robust criterion is presented for computing optimal dynamic inputs for experiments aiming at improving parameter precision. The criterion is based on the information matrix derived for dynamic systems. The design problem is cast as an optimal control problem, with a max-min optimisation objective for design robustness. A typical semi-continuous bioreactor application is presented.